Fatty acid esters of riboflavin



United States Patent FATTY ACH) ESTERS OF RIBOFLAVIN Kunio Yagi, 30l-chome, Meigetsu-cho, Shower-kn, Nagoya-shi, Japan No Drawing. FiledApr. 10, 1962, Ser. No. 186,327 Claims priority, application Japan, Jan.14, 1960,

1 Claim. (Cl. 260211.3)

The present application is a continuation-in-part of ap-. I

plication Serial No. 78,668, filed December 27, 1960, now abandoned.

This invention relates to fatty acid esters'of riboflavin and to aprocess for preparing same.

It has been well known that riboflavin is a slightly water-solublevitamin and, therefore, has some inconveniences for food-enrichment orpharmaceutical fields.

According to this invention 1 have found that riboflavin may be renderedfat-soluble by esterification with fatty acids. A fatty acid ester ofriboflavin can be prepared by reacting riboflavin with an alkanoylhalide having 4 to 20 carbon atoms. The reaction may be carried out inan inert solvent, such as benzene, chloroform or other halogenatedhydrocarbons, in the presence of an acid acceptor such as pyridine.Since riboflavin has four free hydroxy groups in the molecule, it ispreferable to use more than four moles of the alkanoyl halide per moleof riboflavin. As an alkanoyl halide, I prefer to use those having 4 to20 carbon atoms such as butyryl, hexanoyl, octanoyl, capryl, lauroyl,myristoyl, palmitoyl and stearoyl chloride or bromide.

The process of the present invention also includes the use ofcorresponding acid anhydride instead of said acid halide. After thereactants have been heated for the desired length of time, theriboflavin ester is recovered by evaporation of the solvent underreduced pressure or by pouring the reaction mixture into inert solventsin which the riboflavin ester is insoluble.

Among the fatty acid esters, those having lower alkanoyl moiety can beconveniently prepared by directly reacting riboflavin with the alkanoicanhydride in the presence of concentrated perchloric acid. For example,the butyric ester of riboflavin can be obtained in high yield by addinga small amount of 60% perchloric acid into a mixture of riboflavin andbutyric anhydride with stirring at room temperature and then pouring thereaction mixture into water.

The fatty acid esters of riboflavin thus prepared may be recovered andpurified by recrystallization from ether, benzene or ethanol. Theriboflavin esters of this inven tion are substantially tasteless andodorless, and are quite soluble in many conventional solvents, such asethanol, pyridine, benzene, chloroform, or neutral fats, in each ofwhich riboflavin itself is almost insoluble.

The riboflavin esters of this invention have the following structuralformula:

3,189,598 Patented 'June 15, 1965 wherein RCO stands for a straightchain alkanoyl grou containing from 4 to 20 carbon atoms.-

It has been found that the compounds of this invention possess vitamin Bactivity. When administered orally, the compounds are easily hydrolyzedwith human digestive juices into riboflavin and corresponding fattyacids, and effect vitamin B activity. It has been found that thecompounds are'hydrolyzed with neither saliva nor gastric juices, butwith duodenal juice. Among many of the compounds riboflavintetrabutyrate is most easily hydrolyzed. This can be illustrated by thetest described below.

Each of the substrates was suspended in a mixture'of 0.3 ml. of 0.1 Mphosphoric acid-buffered solution (pH 8.5) and 0.2 ml. of human duodenaljuice at a concentration of 1x10- M. The substrate solutions were incu-'bated for 30 minutes at 37 C. and the degradation rate of each of thesubstrates was assayed by the lumiflavin fluorescence method (Yagi, J.Biochem., vol. 43, page It has also been found that the compounds of theinvention show vitamin B activity on dietary scurvy using male albinorats. Among the compounds, riboflavin tetrabutyrate increases the bodyweight of rats as much as free riboflavin and it preventsariboflavinosis remarkably. However, others were found to have littlegrowth-promoting activity.

It has also been found that when the compounds of this invention areadministered to the living body by intramusclar injection, the bloodlevel of total riboflavin rises more slowly, and remains at an effectivelevel for a longer period than with free riboflavimj- Furthermore, theadministered riboflavin is substantially excreted in urine for a longerperiod of time than is free riboflavin. For example, when 5 mg. of freeriboflavin is administered to a rabbit, weighting about 3 kg., byintramuscular injection, the blood level of total riboflavin reaches apeak within 30 to 60 minutes and a substantial quantity of the givenriboflavin is excreted in urine within a few hours. However, in the caseof riboflavin tetrabutyrate, the blood level of total riboflavin reachesa peak on more than 12 hours after the injection, and 30 to 40% of thegiven riboflavin is excreted in urine within 24 hours.

From the foregoing, it is apparent that the compounds of this inventionare useful in nutritional and medical fields, and sometimes moreadvantageously than free riboflavin. The invention is furtherillustrated by the following examples.

Example 1 1 g. of riboflavin was suspended in 300 ml. of a 1:1 mixtureof pyridine-chloroform. Into this suspension, 15 g. of palmitoylchloride in 50 ml. of chloroform was dripped at 0 C. with vigorousstirring within about 1 hour. The mixture was stirred for an additional14- r H, 10.61 N, 4.21..

. yellow oil.

i The solution was allowed to stand overnight .in' a refrigerator,whereby riboflavin tetrapalmitate crystallized out.

7 Example) L1 4;; er riboflavin was s'uspended in '300 m1. of 11-1mixture of pyridine-chloroform. Into this suspension;

10 gr of capryl chloride in 50 ml; of chloroform was dripped at C.withvigorous stirring within about 1 Yield, 2 g yellow crystals meltingat 78. C.. Y after recrystallizationfrom absolute ethanol;

.:iAnalysiS. Calcu1ated C81H146016N4 C; .Found: C, 72.93; H, -10.73,; N,4.41;

: lized out.

hour.= QThe mixture was stirred; for an additional "14 hours at 33 ,C;The reaction mixture was'coneentrated to dryness under reduced pressureand the-residue was treated with 100 ml. of 9 0% pyridine at70 C. for Iminutes in order :to decornpose'the remaining capryl chloride intocapric acid. After-the solution was con-' centrated to dryness, theresidue was dissolved in a small amount of ether.

The elua'te was condensed to remove pyridine whereby riboflavintetracaprate .was obtained. Yield, 1.5 i g.

f 1 offribofiavin was suspended in 300 nila'of 1:1 I rnixtureofpyridine-chloroform. Into this suspension,

5.6 goof butyryl chloride in ml. of chloroform: was

, dripped. at..03 .C. 1 with vigorous stirring within about 1 hour. Themixture was stirred for an-additional '14: hours. at "38 C. i The"reaction mixture was concen trated to dryness under reduced pressureand the residue was treated with mlaof 90% pyridine at 70 C.

fo'r'QOminutes in ordertoidecomposelthe remaining hutyryl chloride intobutyrici acid. 7 After the solution was concentrated to dryness, a smallamount of saturated. solution of sodium bicarbonate in water was addedto the residue and the mixture was. extracted with V The solution waspoured into a column of Florisil (an activated mangesium silicate): Thecolumn was washed with a small amount of ether and ethanol successively,and then eluted with pyridine;

" dride. 25

ether. 7

ether. The extract was dried with sodiumisulfate and then ether wasremoved; The'residue was dissolved in ether and insoluble matter wasfiltered off. The filtrate was condensed, to dryness and the residue wastreated with petroleum ether on a water-bath, and then filtered. Thefilter cake was dissolved in' ether and a major part of the etherwa'siremoved b'yevapora tiona The residue 1 'was cooled, whereby"riboflavin tetrabuty'rate crystal- Yield, 1' -g., yellowish orangecrystals meltafter recrystallization irom j ing at 145 to 147 C.

, Example 4.

' 1 g. of riboflavin was added to elO'mlliof butyricfan hydride; Themixture was heated 5011 an oilbath at C. to C. 'for '6hoi1rs.'The'resultantsolution was concentrated under reduced pressure.Ribofla'vin f tetrabutyrate was crystallized from theresidue" using;

Yield, 1 g., yellowish crystals meltether as a solvent. ing at C. to 147C.

Example' 5 1 of riboflavin was added to 10 inl. of butyric anhy- To themixture was added 05ml. of 60% aque-' ous solution of perchloric acidat. room temperature with .vigorous stirringi The reaction mixture was Vpoured into 50 nil. of water. After the orangefyellowr.

ifsh layer was concentrated under reduced pressure, ribo flavintetrabutyrate was crystallized from ether. g. of riboflavintetrabutyratemelting at 145 C. to 147' V Cewas obtained. 7

'Iclaimy "V Riboflavin 21314? ,5-tetrabutyrate.

I f 'Refe rences Qitedbythe Enaminer V V 'UNITED SVTATES'PA'VTENTS.52,825,729

LEWIs GOTT S Prir nmyExaminer;

3/58;Peterin"g et al. 260 2113 2,970,995" 2/61 .Wheeler. "26O-'- 21V1.37

